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Breakthrough Articles

breakthrough articles

Breakthrough Articles present high-impact studies answering long-standing questions in the field of nucleic acids research and/or opening up new areas and mechanistic hypotheses for investigation. These articles are chosen by the Editors on the recommendation of Editorial Board Members and Referees. Articles are accompanied by a brief synopsis explaining the findings of the paper and where they fit in the broader context of nucleic acids research. They represent the very best papers published at NAR.

Read the full editorial 'Breakthrough Articles: Putting science first'.

2014 Breakthrough Articles available here

Biogenesis and growth phase-dependent alteration of 5-methoxycarbonylmethoxyuridine in tRNA anticodons

Amin Yusuke Sakai, Kenjyo Miyauchi, Satoshi Kimura and Tsutomu Suzuki

A post-transcriptional modification, 5-methoxycarbonylmethoxyuridine (mcmo5U), is present in the anticodon of several bacterial tRNAs. However, biogenesis and function of the terminal methyl group of mcmo5U remained unidentified for more than 35 years. In this study, using precise mass spec analysis, the authors found that mcmo5U was present in four tRNA species responsible for NCN codons. They identified an AdoMet-dependent methyltransferase, CmoM, responsible for mcmo5U formation. A reporter assay revealed that terminal methylation of mcmo5U contributes to the decoding ability of tRNAAla1. Growth phase–dependent alteration of mcmo5U in tRNAPro3, indicates a novel mechanism of translational control mediated by the regulatory decoding efficiency.
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Automated physics-based design of synthetic riboswitches from diverse RNA aptamers

Amin Espah Borujeni, Dennis M. Mishler, Jingzhi Wang, Walker Huso, and Howard M. Salis

Riboswitches are shape-changing regulatory RNAs that bind chemicals and regulate gene expression, directly coupling sensing to cellular actuation. The physical mechanisms that govern a riboswitch’s function remain poorly understood, particularly when changing its ligand-binding aptamer domain. This study presents a model of the physics governing riboswitch function, which entails modeling the ligand-induced RNA conformational changes that directly affect the thermodynamics of self-assembly controlling biological function, and validates the model by designing and characterizing a large number of synthetic riboswitches, using 6 different RNA aptamers, that bind to their respective ligands.
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The casposon-encoded Cas1 protein from Aciduliprofundum boonei is a DNA integrase that generates target site duplications

Alison B. Hickman and Fred Dyda

Many bacteria and archaea have an adaptive immune system called CRISPR-Cas. The CRISPR locus is where DNA sequences from foreign DNA such as bacteriophages or plasmids are stored in the form of an array of short "spacers" that alternate with a palindromic repeat sequence. This record of foreign DNA can be used by CRISPR-associated (Cas) proteins to destroy invading DNA if it tries to re-infect the cell. Insertion of foreign DNA spacers requires two Cas proteins, Cas1 and Cas2. Recently, genes encoding Cas1 homologs have also been identified within putative DNA transposons called casposons. Here, the authors demonstrate that the casposon-associated Cas1 protein from the archaeon Aciduliprofundum boonei is an active DNA integrase with properties that distinguish it from CRISPR-associated Cas1 proteins. Understanding how Cas1 proteins have evolved different biochemical properties may provide insight into the relationships between mobile DNA elements and the evolution of adaptive immune systems.
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Methylation of histone H3 lysine 9 occurs during translation

Rivera C, Saavedra F, Alvarez F, Díaz-Celis C, Ugalde V, Li J, Forné I, Gurard-Levin ZA, Almouzni G, Imhof A, Loyola A.

Post-translational modifications of histones are key contributors to chromatin function. Thus, how these marks are established and by which enzymes are important questions to address. In this study the authors show that H3K9 methylation occurs during the translation of histone H3 by the methyltransferase SetDB1. This is a key mark in the establishment of heterochromatin, a chromatin domain important for gene silencing. Importantly, this is the first histone modification shown to occur during translation, supporting the critical nature of this mark for genome integrity.
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Structure of a human translation termination complex

Matheisl S, Berninghausen O, Becker T, Beckmann R.

In this study, the authors used the termination stalling peptide from human cytomegalovirus (hCMV) to stabilize a pre-termination complex containing a peptidyl-tRNA, the release factor eRF1 and a UAA stop codon in the A-site of the human ribosome. The structure sheds light on the molecular mechanism by which eRF1 can recognize eukaryotic stop codons: in contrast to sense codon recognition and stop codon recognition in bacteria, decoding of the eukaryotic UAA(A) in eukaryotes occurs in an unexpected U-turn geometry. This finding rationalizes a great deal of previously reported biochemical and mutational data, and also illustrates how eRF1 monitors geometry, rather than probing the identity of individual bases, in order to recognize three different stop codons while discriminating against similar sense codons. Translation termination is a fundamental process in every eukaryotic cell. The data in this study not only provides a structural basis for a mechanistic understanding of this process but highlights the substantial mechanistic differences between eukaryotic and prokaryotic translational control and termination.
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FDF-PAGE: a powerful technique revealing previously undetected small RNAs sequestered by complementary transcripts

Harris CJ, Molnar A, Müller SY, Baulcombe DC.

Since their discovery, small RNAs have been found to play a role in an increasingly wide range of cellular processes. Their central mechanism relies on hybridisation with complementary nucleic acid targets. Here, the authors demonstrate that current techniques for small RNA detection are biased by the presence of complementary nucleic acids. They present a method, FDF-PAGE, to overcome this effect. FDF-PAGE reveals key differences in endogenous small RNA profiles and provides insight into viral small RNA biogenesis. This improved technique should be of interest to scientists involved in all fields of small RNA research.
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gkv274A new transcription factor for mitosis: in Schizosaccharomyces pombe, the RFX transcription factor Sak1 works with forkhead factors to regulate mitotic expression

Garg A, Futcher B, Leatherwood J.

In this study, the authors have discovered a major new transcription factor essential in controlling mitotic gene expression in S. pombe. It belongs to the RFX family of transcription factors which have not been previously implicated in cell cycle regulation. Using genome wide techniques such as ChIP-Seq and microarray gene expression analysis, the authors have teased out its function as well as the function of the other major mitotic transcription factors required for mitosis in S. pombe. Cell cycle transcriptional regulation is of general interest to a wide readership, because for instance cancer is a disease of an aberrant cell cycle.
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gkv262Targeting chromatin binding regulation of constitutively active AR variants to overcome prostate cancer resistance to endocrine-based therapies

Chan SC, Selth LA, Li Y, Nyquist MD, Miao L, Bradner JE, Raj GV, Tilley WD, Dehm SM.

Understanding and overcoming mechanisms of resistance to second-generation endocrine-targeted androgen receptor (AR) inhibitors are arguably the most urgent priorities in the field of prostate cancer research. This article demonstrates that AR variants (AR-Vs) arising from alterations in AR mRNA splicing broadly recapitulate chromatin binding events that are otherwise suppressed during endocrine-targeted therapy. Additionally, this work reveals that the bromodomain inhibitor JQ1 down-regulates AR-V expression, which leads to impaired AR-V chromatin binding, impaired transcriptional activation of target genes, and impaired growth of AR-V-driven prostate cancer cells in vitro and in vivo.
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gkv302Telomere-associated proteins add deoxynucleotides to terminal proteins during replication of the telomeres of linear chromosomes and plasmids in Streptomyces

Yang CC, Tseng SM, Chen CW

The linear chromosomes and plasmids in soil bacteria Streptomyces are capped by covalently-linked terminal proteins (TPs), which are primers for initiation of DNA synthesis at the ends of the DNA. Specific proteins, named telomere-binding protein (Tap), bind the TPs and the ends of DNA, and are necessary for replication. In this study, the authors demonstrate that Taps can perform the priming reaction in the absence of a DNA polymerase. These proteins represent a new class of DNA synthesis enzymes.
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gkv243Next-generation sequencing reveals the biological significance of the N2,3-ethenoguanine lesion in vivo

Chang SC, Fedeles BI, Wu J, Delaney JC, Li D, Zhao L, Christov PP, Yau E, Singh V, Jost M, Drennan CL, Marnett LJ, Rizzo CJ, Levine SS, Guengerich FP, Essigmann JM

This study illustrates effective use of a high-throughput sequencing approach to multiplex the investigation of the genotoxicity, mutagenicity and repair of multiple DNA lesions in multiple cell strains. The authors applied this improved sequencing approach to investigate the ethenoguanine lesions, which are formed after vinyl chloride exposure or oxidative stress. The results not only show that this high-throughput approach can yield accurate and precise measurements, but also highlight the significance of the N2,3-ethenoguanine lesion. This lesion potently induces the same type of mutation found in vinyl chloride-induced tumors. The work shows that it is poorly repaired compared to other etheno lesions.
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gkv220H3K4 monomethylation dictates nucleosome dynamics and chromatin remodeling at stress-responsive genes

Nadal-Ribelles M, Mas G, Millán-Zambrano G, Solé C, Ammerer G, Chávez S, Posas F, de Nadal E.

In response to extracellular stimuli, stress-activated protein kinases (SAPK) modulate gene expression to maximize cell survival to stress. The yeast Hog1 SAPK has been instrumental to understand gene expression regulation in response to environmental stimuli. Here, the authors present genetic and biochemical evidence that expression of stress-responsive genes is controlled through two independent remodeling mechanisms: RSC in the presence of monomethylated H3K4 and Swr1 in the absence of H3K4 monomethylation. These results point to a novel role for H3K4 monomethylation in dictating the specificity of chromatin remodeling, which functions as an additional layer of transcriptional control of stress-responsive genes.
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gkv176AKT phosphorylates H3-threonine 45 to facilitate termination of gene transcription in response to DNA damage

Lee JH, Kang BH, Jang H, Kim TW, Choi J, Kwak S, Han J, Cho EJ, Youn HD.

AKT1 silencing decreases cell survival on DNA damage but its detailed molecular mechanism has not been identified so far. In this study, the authors have identified AKT1 as a novel histone kinase, phosphorylating H3-T45 under DNA damaging conditions. They discovered that H3-T45 phosphorylation on transcription termination site regulates 3’ end processing of DNA damage-responsive genes to facilitate overall transcription efficiency, providing an unexpected function of AKT1 in transcription termination.
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gkv195 Integrating motif, DNA accessibility and gene expression data to build regulatory maps in an organism

Blatti C, Kazemian M, Wolfe S, Brodsky M, Sinha S.

In this work, the authors demonstrate how understanding of cell type specific regulatory networks and elements can be obtained from integrating gene expression and chromatin accessibility data with computationally predicted profiles of transcription factor (TF) binding. With a comprehensive collection of TF binding motifs, they apply this novel procedure to explore poorly characterized gene expression domains of fruitfly embryonic development. The results show improvement over methods that rely on experimentally determined TF binding from available, but limited ChIP data. The approach to decode gene regulation presented here is most useful to biologists who study non-model organisms or specific cell types in which conducting comprehensive experimental assays is technically or financially infeasible.
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gkv172Crystal structure of Hop2–Mnd1 and mechanistic insights into its role in meiotic recombination

Kang HA, Shin HC, Kalantzi AS, Toseland CP, Kim HM, Gruber S, Dal Peraro M, Oh BH.

During meiotic cell division, the Hop2–Mnd1 heterodimer plays an important role in Dmc1 nucleofilament-mediated homologous recombination, which results in invasion of Dmc1-coated single-stranded DNA into double-stranded DNA between homologous chromosome pairs. An X-ray crystallographic study reveals that Hop2–Mnd1 adopts an elongated crescent-like structure containing a double-stranded DNA-binding domain at one end and a Dmc1 nucleofilament-binding domain at the other end. Structure-based studies suggest that Hop2–Mnd1 perturbs base pairing of recipient double-stranded DNA, and that it juxtaposes perturbed DNA close to invading single-stranded DNA through its bipartite structure to promote search for complementary DNA sequence between the two DNAs.
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gkv078CarD stabilizes mycobacterial open complexes via a two-tiered kinetic mechanism

Rammohan J, Ruiz Manzano A, Garner AL, Stallings CL, Galburt EA.

Individual bacterial species use specialized strategies and factors to regulate gene expression. The study describes the kinetic mechanism of CarD, an essential transcription factor in many bacteria including Mycobacterium tuberculosis, but not even present in others (i.e. Escherichia coli). The work also reveals a fundamental difference between the RNA polymerases from a CarD-containing and a CarD-acking bacteria and rationalizes the essentiality of CarD in species where it exists. This highlights the importance of studying non-model systems, even within the context of a highly conserved process like transcription, and advances our understanding of the physiology of this important pathogen.
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gkv009Sequence-specific cleavage of dsRNA by Mini-III RNase

Głów D, Pianka D, Sulej AA, Kozłowski LP, Czarnecka J, Chojnowski G, Skowronek KJ, Bujnicki JM.

Deoxyribonucleases that cut double stranded DNA, such as restriction enzymes, have contributed to the revolution in biotechnology. However, thus far no anologous enzymes were available for cleavage of double stranded RNA. Here, the authors present evidence for a sequence-dependent cleavage of long dsRNA by ribonuclease Mini-III from Bacillus subtilis (BsMiniIII), a member of RNase III superfamily. These results suggest that BsMiniIII may serve as a prototype of a sequence-specific dsRNase that could possibly be developed towards a “restriction enzyme for RNA.”
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gku1395A systematic survey of the Cys2His2 zinc finger DNA-binding landscape

Persikov AV, Wetzel JL, Rowland EF, Oakes BL, Xu DJ, Singh M, Noyes MB.

Cys2-His2 zinc fingers (C2H2-ZFs) are the most abundant class of DNA-binding proteins in higher organisms, and have been implicated in a wide range of biological processes. Furthermore, C2H2-ZFs can be engineered to specifically target genomic regions for therapeutic or other applications. Nevertheless, our understanding of how C2H2-ZFs specify their targets remains incomplete. To help bridge this gap in fundamental knowledge, the authors have performed a comprehensive series of screens to detect and quantify C2H2-ZF – DNA interactions. This work provides new insights into the complex DNA-binding landscapes of C2H2-ZFs, and serves as a valuable resource for further investigation of this important domain.
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